The proposed research is directed toward a combined biochemical and genetic analysis of the molecular processes involved in DNA repair in eukaryotes. Drosophila melanogaster provides a multicellular model system for the study of the genetic control of these processes and their relationship to mutation induction by physical and chemical agents encountered in the environment. In analogy with microbial systems, an array of mutagen-sensitive strains of Drosophila have been isolated to provide mutants defective in various DNA repair functions (Smith 1973, 1976; Boyd et al, 1976a). The interrelationships among these mutant loci with respect to mutagen cross sensitivity, repair proficiency, double mutant interactions, and proficiency in mitotic and meiotic functions provide information necessary for identification and ordering of the intermediate steps in the complex pathways involved in repair of damaged DNA. This information will be used to select a restricted number of mutants affecting particular repair processes for detailed biochemical analysis designed to detect altered gene products. Three approaches to the biochemical examination of these mutants are outlined: Assays will be performed to determine excision repair proficiency of selected mutants strains. A two-dimensional gel electrophoresis technique will be utilized to detect altered or missing gene products in the mutant strains. Fractionation of cellular proteins by DNA affinity techniques prior to electrophoresis will be used to enrich for DNA repair associated activities. Both autoradiographic and specific enzymatic assay methods will be used to localize proteins on gels. Specific repair associated enzymes will be isolated from mutant strains exhibiting phenotypes which strongly implicate a particular defect in DNA repair and examined for altered in vitro activity.